Biology subjects

MacPhee, C. E.

Publications and source records attributed to MacPhee, C. E..

7 recordsLinked to original sources

bioRxiv2025

Regulatory Rewiring Drives Intraspecies Competition in Bacillus subtilis

Intraspecies interactions shapes microbial community structure and evolution, yet the mechanisms determining competitive outcomes among closely related strains remain unclear. The soil bacterium Bacillus subtilis is a model for microbial social interactions, where quorum-sensing systems regulate cooperation and antagonism. Here, we take a multifaceted approach to dissect the role of quorum-sensing regulation in competitive fitness. Isolate NCIB 3610 carries a signal unresponsive RapP-PhrP module that alters quorum-sensing control and promotes faster growth. Modelling and mutant analysis demonstrate that the small differences in growth rate conferred by RapP-PhrP3610 are sufficient to drive competitive exclusion. The importance of quorum sensing control is further exemplified by experimental evolution of distinct wild isolates, which revealed recurrent mutations in the sensor kinase comP, which phenocopy complete comP or comA deletions and confer a growth-linked competitive advantage. Key quorum sensing mechanisms are abandoned even in structured microbial communities, where it might be expected that communal traits are favoured. Furthermore, a phylogenomic survey of 370 B. subtilis genomes identified disruptive comP mutations in [~]16% of isolates. However, growth rate alone does not explain all interaction outcomes as even isogenic strains with equivalent doubling times differ in competitiveness. Transcriptomic profiling and validation experiments implicated a type VII secretion system toxin as an additional effector. These findings reveal that disruption of quorum-sensing pathways, whether naturally or through selection, provides a rapid route to competitive advantage, highlighting a fundamental trade-off between communal signalling and individual fitness in microbial populations. ImportanceMicrobial competition and cooperation are key in shaping the structure and evolution of microbial communities. Our study on Bacillus subtilis, a model for microbial social interactions, reveals how alterations in cell-cell communication can enhance competitive fitness. We show, through a combination of modelling, mutational analysis, and experimental evolution, that certain strains of B. subtilis gain competitive advantage by disrupting their quorum sensing pathways, which leads to faster growth and enhanced competitiveness. Such mutations are prevalent in [~]16% of analysed genomes, underscoring their widespread evolutionary benefit under competitive conditions. Our research highlights the delicate balance between individual success and community cooperation in microbes, providing insights into microbial behaviour and potential applications in modifying microbial ecosystems.

microbiology↗

bioRxiv2021

Lipid-induced polymorphic amyloid fibrils formation by α-synuclein.

Many proteins that self-assemble into amyloid and amyloid-like fibres can adopt polymorphic forms. These forms have been observed both in vitro and in vivo and can arise through variations in the steric-zipper interactions between {beta}-sheets, variations in the arrangements between protofilaments, and differences in the number of protofilaments that make up a given fibre class. Different polymorphs which arise from the same precursor molecule not only exhibit different levels of toxicity, but importantly can contribute to different disease conditions. In this work, we show that in the presence of 1,2-dimyristoyl-sn-glycero-3-phospho-L-serine, a highly abundant lipid in the plasma membrane of neurons, the aggregation of -synuclein is markedly accelerated and yields a diversity of polymorphic forms under identical experimental conditions. This morphological diversity includes thin and curly amyloid fibrils, helical and twisted ribbons, nanotubes and flat sheets. TEM analysis of fibrils sampled from the early stage of the growth phase shows the presence of helical and twisted ribbons, indicating that these morphological variants form at the early stages of aggregation. Total internal reflection fluorescence microscopy (TIRFM) indicated the presence of lipids collocated with the mature fibrils. This finding has important implication as the presence of -synuclein with co-localized high lipid content has been reported in Lewy bodies, the pathological hallmark of Parkinsons disease and other synucleinopathies. Thus, the present study demonstrates that an interface, such as that provided by a lipid membrane, can not only modulate the kinetics of -synuclein amyloid aggregation but also plays an important role in the formation of morphological variants by incorporating lipid molecules in the process of amyloid fibril formation.

biophysics↗

bioRxiv2021

Founder cell configuration drives competitive outcome within colony biofilms

Bacteria typically form dense communities called biofilms, where cells are embedded in a self-produced extracellular matrix. Competitive interactions between strains within the biofilm context are studied due to their potential applications in biological, medical, and industrial systems. Combining mathematical modelling with experimental assays, we reveal that the spatial structure and the competitive dynamics within biofilms are significantly affected by the location and density of founder cells. Using an isogenic pair of Bacillus subtilis strains, we show that the observed spatial structure and relative strain biomass in a mature biofilm can be mapped directly to the locations of founder cells. Moreover, we define a predictor of competitive outcome that accurately forecasts relative abundance of strains based solely on the founder cells access to free space. Consequently, we reveal that variability of competitive outcome in biofilms inoculated at low founder density is a natural consequence of the random positioning of founding cells in the inoculum. Extending our study to non-isogenic strain pairs of B. subtilis, we show that even for strains with different antagonistic strengths, a race for space remains the dominant mode of competition in biofilms inoculated at low founder densities. Our results highlight the importance of spatial dynamics on competitive interactions within biofilms and hence to related applications.

microbiology↗

bioRxiv2020

Density and temperature controlled fluid extraction in a bacterial biofilm is determined by poly-γ-glutamic acid production

A hallmark of microbial biofilms is the self-production of extracellular matrix that encases the cells resident within the community. The matrix provides protection from the environment, while spatial heterogeneity of expression influences the structural morphology and colony spreading dynamics. Bacillus subtilis is a model bacterial system used to uncover the regulatory pathways and key building blocks required for biofilm growth and development. Previous reports have suggested that poly-{gamma}-glutamic acid (PGA) production is suppressed during biofilm formation and does not play a major role in biofilm morphology of the undomesticated isolate NCIB 3610. In this work we report on the observation of multiple travelling fronts that develop during the early stage of B. subtilis colony biofilm formation. We find the emergence of a highly motile population of bacteria that is facilitated by the extraction of fluid from the underlying agar substrate. Motility develops behind a moving front of fluid that propagates from the boundary of the biofilm towards the interior. The extent of proliferation is strongly modulated by the presence of extracellular polysaccharides (EPS). We trace the origin of this moving front of fluid to the production of PGA. We find that PGA production is correlated with higher temperatures, resulting in a mature biofilm morphology that is distinct from the biofilm architecture typically associated with B. subtilis. Our results suggest that B. subtilis NCIB 3610 produces distinct biofilm matrices in response to environmental conditions.

microbiology↗

bioRxiv2017

Formation of functional, non-amyloidogenic fibres by recombinant Bacillus subtilis TasA

Bacterial biofilms are communities of microbial cells encased within a self-produced polymeric matrix. In the Bacillus subtilis biofilm matrix the extracellular fibres of TasA are essential. Here a recombinant expression system allows interrogation of TasA, revealing that monomeric and fibre forms of TasA have identical secondary structure, suggesting that fibrous TasA is a linear assembly of globular units. Recombinant TasA fibres form spontaneously, and share the biological activity of TasA fibres extracted from B. subtilis, whereas a TasA variant restricted to a monomeric form is inactive and subjected to extracellular proteolysis. The biophysical properties of both native and recombinant TasA fibres indicate that they are not functional amyloid-like fibres. A gel formed by TasA fibres can recover after physical shear force, suggesting that the biofilm matrix is not static and that these properties may enable B. subtilis to remodel its local environment in response to external cues. Using recombinant fibres formed by TasA orthologues we uncover species variability in the ability of heterologous fibres to cross-complement the B. subtilis tasA deletion. These findings are indicative of specificity in the biophysical requirements of the TasA fibres across different species and/or reflect the precise molecular interactions needed for biofilm matrix assembly.\n\nContributionsConceived and designed the experiments: CE, EE, RG, CEM, RJM, MS, NSW; Performed the experiments: KB, LC, CE, EE, PKF, RG, CEM, RJM, MS, TS; Contributed new analytical tools: CE, EE, RG, TS; Analysed the data: CE, EE, CEM, RJM, MS, LCS, NSW; Wrote the paper: EE, RJM, CEM, MS, NSW.

microbiology↗

bioRxiv2017

Bifunctionality Of A Biofilm Matrix Protein Controlled By Redox State

Biofilms are communities of microbial cells that are encapsulated within a self-produced polymeric matrix. The matrix is critical to the success of biofilms in diverse habitats, but despite this many details of the composition, structure, and function remain enigmatic. Biofilms formed by the Gram-positive bacterium Bacillus subtilis depend on the production of the secreted film-forming protein BslA. Here we show that a gradient of electron acceptor availability through the depth of the biofilm gives rise to two distinct functional roles for BslA and that these can be genetically separated through targeted amino acid substitutions. We establish that monomeric BslA is necessary and sufficient to give rise to complex biofilm architecture, while dimerization of BslA is required to render the community hydrophobic. Dimerization of BslA, mediated by disulfide bond formation, depends on two conserved cysteine residues located in the C-terminal region. Our findings demonstrate that bacteria have evolved multiple uses for limited elements in the matrix, allowing for alternative responses in a complex, changing environment.\n\nSignificanceThe biofilm matrix is a critical target in the hunt for novel strategies to destabilise or stabilise biofilms. Knowledge of the processes controlling matrix assembly is therefore an essential prerequisite to exploitation. Here we highlight that the complexity of the biofilm matrix is even higher than anticipated with one matrix component making two independent functional contributions to the community. The influence the protein exerts is dependent on the local environmental properties, providing another dimension to consider during analysis. These findings add to the evidence that bacteria can evolve multifunctional uses for the extracellular matrix components.

microbiology↗

bioRxiv2016

Evolutionary variations in the biofilm-associated protein BslA from the genus Bacillus

BslA is a protein secreted by Bacillus subtilis which forms a hydrophobic film that coats the biofilm surface and renders it water-repellent. We have characterised three orthologues of BslA from Bacillus amyloliquefaciens, Bacillus licheniformis and Bacillus pumilus as well as a paralogue from B. subtilis called YweA. We find that the three orthologous proteins can substitute for BslA in B. subtilis and confer a degree of protection, whereas YweA cannot. The degree to which the proteins functionally substitute for native BslA correlates with their in vitro biophysical properties. Our results demonstrate the use of naturally-evolved variants to provide a framework for teasing out the molecular basis of interfacial self-assembly.

microbiology↗